The present invention relates to a device for separating desired biological material(s) from a mixture of biological materials that are in solution or suspended in a liquid. In the past, many devices have been developed to separate mixtures of biological materials from one another. Such biological materials include, DNA, DNA segments, proteins, nucleic acids, other cellular components etc. The separation can be done on the basis of molecular size, molecular weight, affinity for attraction to various filter materials, e.g., adsorptive capacity, etc.
To achieve separation, the materials are preferably dissolved in a solvent to form a solution and the solution is then passed through a separation column as is known in the art. The packing (filter material) in the column and various chemicals that can be added to the column will determine what is collected in the column and what passes through the column to form an eluate that is collected. For example, the column and process can allow small molecules to be separated out in the column and the large molecules to pass through as an eluate. Conversely, the large molecules could be collected in the column and the small molecules could pass through the column in the eluate to be collected for further processing. These separation techniques are well known in the art and the general techniques are described in Molecular Cloning: A Laboratory Manual 2nd ed. Vol""s 1, 2 and 3 by J. Sambrook, E. F., Fritsch and T. Maniatis, published by Cold Spring Harbor Laboratory, 1989 which are incorporated herein by reference.
The devices used in such processes are generally complicated in structure and operation. One such device is described in U.S. Pat. No. 5,824,224 and is usable to extract and purify DNA. This device handles multiple samples simultaneously and is complicated in structure and operation requiring that the various solutions pass through several stages and containers. For example, the disclosed device uses a plurality of eluate collection chambers requiring a separate housing to apply the vacuum simultaneously to the collection chambers. Such an apparatus is not adapted for effective use when a small quantity of samples or a single sample needs to be processed.
U.S. Pat. No. 5,725,763 describes a simpler filter device that utilizes vacuum to induce a liquid to flow thru a side inlet from a storage container thru a filter to a collection container. The relatively short filter housing sits on top of the collection container with a seal therebetween and appears to be held in place by vacuum. The collection container appears to be a standard small mouth container of substantial volume and further appears to be adapted solely for collection and storage. Further, the connection between the container and the filter is not secure and therefore, there is a risk that the device could be unstable in operation.
The present invention provides an improved and simpler device and method for separating biological materials from one another and is capable of using standard separation columns and techniques. The invention includes a separation column connected in flow communication with a reservoir having a collection chamber adapted for collection of filtrate or eluate. The container containing eluate that is not of interest can be discarded and a clean container installed to collect the next eluate or filtrate. This simplifies the overall separation process and reduces expense and the risk of contamination. The reservoir can also include a concentration zone at the bottom of the collection chamber. The concentration zone has a small volume to height ratio at the bottom of the concentration zone to assist in separating the eluate or filtrate into liquid and solid portions in a centrifuge. The concentration zone is particularly useful when the eluate contains only a small percentage of solid material. A vacuum source is connected to the device to induce flow of liquid thru the separation column and into the reservoir. The invention is also adapted for processing a single sample easily, effectively and economically. The simplicity of the inventive device reduces cost allowing the device to be disposed of after use. Disposability eliminates the need for subsequent equipment clean up and clean up costs.
Among the several objects and features of the present invention may be noted the provision of a device and method that are used to separate a mixture of biological materials and capture the desired biological material for further processing and use; the provision of a method that is effective and economical in separating biological materials; the provision of such a method that reduces clean up after practice, the provision of such a method that can be used on a wide variety of materials; the provision of such a device that is simple in construction and inexpensive to manufacture; the provision of such a device that can be disposed of after use; the provision of such a device that includes a collection container that can be used in the collection of eluate and in subsequent separation steps like centrifugation; the provision of such a device that uses a pressure differential to induce flow of liquid through the separation column into a reservoir for collection in a collection chamber; and the provision of such a device that is simple in construction and effective in operation requiring minimal component parts.
One aspect of the present invention involves the provision of a separation device for separating biological materials. The device includes a reservoir comprising a container and a lid removably secured to the container. The container defines a liquid collection chamber and has an interior bottom surface forming a concentration zone at the bottom of the collection chamber for collecting solids. The concentration zone has a transverse cross sectional area that increases in size from the bottom of the concentration zone to the top of the concentration zone. A first connector is in fluid flow communication with the collection chamber and is adapted for connection to a vacuum source. A separation column is operatively associated with the reservoir and is in fluid flow communication with the collection chamber. The separation column contains packing material operable to separate mixed biological materials. The column has an inlet and an outlet whereby in operation, liquid containing biological materials to be separated is introduced into the column thru the inlet with a portion of the biological material being retained in the column and a portion of the biological material passes thru the column with the liquid and out the outlet for collection in the collection chamber.
The present also involves the provision of a method of separating biological materials from a mixture of biological materials. The method includes preparing a mixture of liquid and biological materials. A portion of the liquid mixture is introduced into a separation column. Flow of a portion of the introduced liquid mixture is induced through the column with the column allowing a portion of the liquid mixture to pass through the separation column. A portion of the liquid mixture passing through the separation column is collected in a collection chamber of a container with the collection chamber having a concave concentration zone in a bottom portion of the chamber. The container with the collected portion of the liquid mixture therein is placed in a centrifuge. The thus collected portion of the liquid mixture is centrifuged in the container whereby certain biological material contained in the liquid is concentrated and collected in the concentration zone. The concentrated biological material is separated from a major portion of the liquid.
Another aspect of the invention includes a method of separating nucleic acids. The method includes making a solution of nucleic acids and solvent. The solution is placed in a separation column having an inlet and an outlet. A vacuum is applied to the column whereby there is a lower pressure at the outlet than at the inlet thereby inducing flow of a portion of the solution out of the column through the outlet to provide a first eluate and the column retains a portion of the nucleic acids. The first eluate is collected in a container and is then removed from the container. A liquid is introduced into the column to render the retained nucleic acids in the column to a flowable condition in the column. A vacuum is applied to the column to induce the retained nucleic acids and liquid to flow out of the column into the container for collection thereof as a second eluate. The second eluate is centrifuged to concentrate at least a portion of the nucleic acids in the second eluate and the concentrated nucleic acids are collected in a concave concentration zone of the container. A major portion of the liquid in the second eluate is separated from the concentrated nucleic acid material and the separated and concentrated nucleic acid material is recovered.
A further aspect of the invention includes the provision of a separation device for separating materials suspended in liquid. The device includes a reservoir comprising a container and a lid removably secured to the container with the container and lid defining an enclosed liquid collection chamber. A first connector is in fluid flow communication with the collection chamber and adapted for connection to a vacuum source. A separation column is mounted on the lid and is in fluid flow communication with the collection chamber. The separation column contains packing material that is operable to separate mixed materials in a liquid. The separation column has an inlet and an outlet whereby in operation, liquid containing materials to be separated is introduced into the column thru the inlet with a portion of the materials being retained in the column and a portion of the materials passing thru the column with the liquid and out the outlet for collection in the collection chamber.
Other objects and features will be in part apparent and in part pointed out hereinafter.